a) Representative flow plots of SPF, germ-free reconstituted and germ-free mice thymi and quantification of the percent of each Treg cell lineage subset within CD4+CD73- thymocytes. b) Representative flow plots and quantification of SPF, germ-free reconstituted and germ free mice spleens showing the percent of CD4+ lymphocytes which are Foxp3+. a,b) Data is representative of 1 experiment, n = 6 SPF mice, 5 germ-free reconstituted mice, and 6 germ-free mice. Data was analyzed by one-way ANOVA with Tukey’s multiple comparisons test. c) Representative flow plots of CD4+CD73- thymocytes from SPF mice or mice with normalized microbial experience (NME) and quantification of the percent within each Treg cell lineage population. Data is representative of 2 experiments, n = 3 SPF mice and 5 NME mice. Data was analyzed by two-sided unpaired t test. All bars represent mean ± SD. *P<0.05, **P<0.005, ns- not significant.